The inducible and broadly expressed NF-kappaB/Rel family of transcription factors plays a pivotal regulatory role in such diverse processes as proliferation, anti-apoptosis and immune and inflammatory responses. Activation of NF-kappaB results from the select phosphorylation and degradation of IkappaB, a cytoplasmic inhibitor of the NF-kappaB/Rel proteins. This phosphorylation is mediated by a pair of highly related IkappaB kinases, IKK1 and IKK2 which reside as hetero- or homo-dimers in multimeric complexes called signalsomes. Despite the ability of both kinases to phosphorylate the IkappaB substrate, targeted disruption of the genes for these enzymes results in dramatically different phenotypes. IKK2 -/- mice die at embryonic day 13 due to massive hepatic destruction, reflecting a lack of the anti-apoptotic effects of NF-kappaB. In contrast the IKK1-/- animals exhibit a hyperplastic, thickened epidermis due to the failure of IKK1-/- keratinocytes to undergo normal terminal differentiation. This phenotype is also observed with in vivo inhibition of NF-kappaB. Intriguingly, activation of NF-kappaB is only slightly impaired in the IKK1-/- mice. We propose a series of experiments designed to delineate the molecular basis for the IKK1-/- skin phenotype. Specifically, we will test the ability of epidermally targeted expression of either kinase proficient or kinase deficient IKK1 transgenes to repair the abnormal skin phenotype (Specific Aim 1). Secondly, we will evaluate the activation of NF- kappaB in response to TNFalpha, IL-1 or UV light in primary reconstituted keratinocyte versus non-reconstituted fibroblast cultures from these transgenic mice (Specific Aim 2). Finally, to specifically test the potential role of IKK2 in epidermal differentiation, we will introduce a kinase deficient IKK2 transgene into the epidermis of normal mice (Specific Aim 3). Together these studies should help elucidate the respective contributions of IKK1/2 heterodimers and IKK1 homodimers in regulating epidermal differentiation.